A Modified Simple and Rapid Reversed Phase High Performance Liquid Chromatographic Method for Quantification of Amitriptyline and Nortriptyline in Plasma

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Abstract
A simple and rapid reversed-phase liquid chromatographic method for the determination of amitriptyline and nortriptyline in plasma is described. Protriptyline was used as internal standard. Plasma samples were extracted from alkaline pH with hexane-isoamylalcohol 98:2 v/v for 60 sec on a vortex apparatus. The organic phase was back extracted with HCl 0.1N and the aqueous phase was injected directly and monitored at 240nm. Samples were chromatographed on a 5 μm Kromasil 100-C8 column (25 cm × 4 mm i.d) using acetonitrile/0.04M KH2PO4 40:60 v/v as the mobile phase. The lower limit of detection was 5 ng/ml for amitriptyline and nortriptyline while the lower limit of quantification was 10 ng/ml for both compounds. Peak height and plasma amitriptyline or nortriptyline concentrations were linearly related from 25 to 300 ng/ml. No potential sources of interference have been identified. The methodology was used to monitor plasma amitriptyline or nortriptyline concentrations in patients receiving therapeutic doses of these drugs as well as for qualitative determination of several tricyclic antidepressants in plasma in cases of emergency toxicology.

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